Method for uptake of a substance into a seed

ABSTRACT

A method of enhancing a growth characteristic of seed by immersing the seed in an aqueous solution including dissolved inert gas and sonicating the seed at a frequency preferably of between about 15 kHz and about 30 kHz and an energy density of between about 1 watt/cm 2  and about 10 watts/cm 2  for a period of between 1 minute and about 15 minutes. The sonicated seed exhibits an enhanced growth characteristic including resistance to pests and growth properties consistent with the introduction of essential nutrients. Plants grown from the treated seeds exhibit improved characteristics.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The invention relates generally to an imbibition process for the uptakeof water or other substances along with dissolved substances into aseed, more specifically, to a method of treating seeds with sound wavesfor the purpose of improving the rate of water and/or substance uptakeinto the seed wherein the substance enhances a growth characteristic ofthe seed or added value of the seed during commercial processing.

2. Background of the Prior Art

Seed dormancy is a unique form of developmental arrest utilized by mostplants to temporally disperse germination and optimize progeny survival.During seed dormancy, moisture content and respiration rate aredramatically lowered. The initial step to break seed dormancy is theuptake of water (imbibition) necessary for respiration and mobilizationof starch reserves required for germination. Imbibition is a biphasicprocess: 1) the physical uptake of water through the seed coat andhydration of the embryo and 2) germination as determined by growth andelongation of the embryonic axis resulting in emergence of the plumuleand radicle. The two phases are separated temporally and seed which hascompleted phase I is said to be “primed seed,” that is, primed for phaseII: germination. Phase I of imbibition is also used in the commercialprocessing of seed, i.e. wet milling fractionation of corn and themalting process for the fermentation of distilled spirits.

Priming of seed by the enhanced imbibing of water is advantageous toplant vigor, e.g. enhanced emergence, growth and yield characteristics.Seed priming also synchronizes the germination of seed resulting in anuniform field of plants that matures simultaneously for maximal yieldsat harvest. In addition to water, seed priming provides access to loadthe seed with nutrients, microorganisms or pest inhibitors to promoteseedling establishment. By adding the molecule to the seed duringimbibition phase I, the molecule or organism can be stored in the primedseed and therefore, be present at planting. The “loading ofmacromolecules” is very efficient in the seed when compared to theaddition of similar molecules to the entire field. An example is theaddition of fertilizer to stimulate root growth and hasten seedlingemergence. The loading of the fertilizer into the seed prior to plantingis more efficacious to the seedling and cost effective to the farmer.Other beneficial molecules to be loaded into seed are hormones such asthe gibberelins/gibberellic acid to promote germination, cytokinins forcell elongation and inhibitors of abscisic acid to promote release fromseed dormancy. Seed cultivars could be customized to specific growingregions by the addition of triazoles (plant growth regulators whichmoderate the effects of drought and high temperatures) or fungicides toinhibit the growth of fungi on seed and seedlings in cool, wet soil orinsecticides to combat insects that attack seedlings such as cornrootworm. In addition to macromolecules, beneficial microorganisms suchas Azospirillum or Rhizobium can be loaded during seed priming as a cropinnoculant.

The commercial fractionation of corn begins with wet milling. Corn is acomplex mixture of starch, protein, oil, water, fiber, minerals,vitamins and pigments. Wet milling is the process of separating the corncomponents into separate, homogenous fractions. In Iowa, approximately20% of the 1 billion bushels of corn harvested each year is wet milled.The wet milling industry and collateral manufacturers represent aprodigious industrial effort. As the wet milling process is constantlyrefined by new technologies, novel by-products can be isolated inindustrial quantities, e.g. ethanol, corn sweeteners, protein peptidesand vitamins C and E. The initial step in wet-milling, steeping, has notbeen altered by technological innovation. Steeping involves soaking theclean and dried corn (<16% water content) in warm water until it hasswollen to 45% hydration. This process takes from 30-50 hr. attemperatures of 120-130° F. During the steeping process, largequantities of water are moved through massive vats of corn in acountercurrent stream. Also during this time, beneficial microorganismssuch as the lactobacteria and Pseudomonas aeruginosa growing in thesteep water aid in the proteolytic cleavage of corn proteins. However,the large volumes of steep water and the time required for hydrationlimit the effectiveness of the bacterial digestion. The digestionby-products are purified from the steep water primarily by evaporativeconcentration.

The malting process is the first step in the fermentation of grain toproduce alcoholic spirits. The quality of the malt (and the resultingfermentation) is dependent upon the synchronous and efficientgermination of the grain. Starches stored in the seed are converted intosugars during early stages of germination. At emergence, germination ishalted and converted sugars are used during fermentation for theproduction of ethanol. Historically, the malting process was a laborintensive task. The grain was spread onto a malting floor, imbibed withwater from overhead sprinklers, and turned by hand daily over the courseof one to two weeks to release trapped heat and gases. At plumuleemergence, the starches have been converted to sugars, the germinatedgrain is kiln dried and ground to form malt. Microbreweries anddistilleries still use variations of this old malting technique toproduce high quality malt. Some distilleries induce uniform germinationby the addition of gibberllic acid (GA) to produce the highest qualityof malt for fermentation such as in single malt scotch distillation. GAis the plant hormone which regulates germination. Malt production ofthis quality is time consuming and expensive.

SUMMARY OF THE INVENTION

The invention consists of an imbibition process for the uptake of waterand/or a beneficial substance into seed. The seed to be treated isimmersed in water that includes molecules capable of enhancing a growthcharacteristic or commercial value of the seed. The seed is exposed tosound energy at frequencies between 15 kHz and 30 kHz for periodsbetween about 1 and 15 minutes. The ultrasonic energy generatescavitational forces by the adiabatic collapse of microbubbles in theliquid medium, particularly those bubbles that collapse at the surfaceof the seed. The effect is substantially enhanced by saturating thewater with a noble gas such as helium or argon, or combinations of inertgases.

Seed treated by this method upon germination exhibits an enhanced growthcharacteristic consistent with exposure to the given substance. Thetreated seed can be dried, stored and germinated at a later date whilemaintaining its enhanced growth characteristics.

A purpose of the invention is to impart upon seeds through an imbibitionprocess an enhanced growth characteristic.

These and other objects of the invention will be made clear to a personof ordinary skill in the art upon a reading and understanding of thisspecification, the associated drawings, and appending claims.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a diagrammatic view of the apparatus for practicing thepresent invention.

FIG. 2 shows a table with data from an experiment conducted with cornseed hybrid sonicated in helium saturated tap water.

FIG. 3 shows a table with data from an experiment conducted with cornseed hybrid sonicated in argon saturated water.

FIG. 4 shows a table with data from an experiment identical to theexperiment that generated the data for the table of FIG. 3, exceptperformed with a different corn seed hybrid.

FIG. 5 shows a table with data from an experiment identical to theexperiment that generated the data for the table of FIG. 2 exceptperformed with argon and helium saturated tap water.

FIG. 6 shows a table with data from an experiment identical to theexperiment that generated the data for the table of FIG. 3 exceptperformed with a different corn seed hybrid.

FIG. 7 shows a table with data from an experiment identical to theexperiment that generated the data for the table of FIG. 2 exceptperformed with argon saturated water.

FIG. 8 shows a table with data from an experiment identical to theexperiment that generated the data for the table of FIG. 2 exceptperformed with tap water.

FIG. 9 shows a table with data from an experiment identical to theexperiment that generated the data for the table of FIG. 2 exceptperformed with boiled double distilled water.

FIG. 10 shows a table with data from a different experiment conductedwith corn seed hybrid sonicated in argon saturated tap water.

FIG. 11 shows a table with data from an experiment identical to theexperiment that generated the data for the table of FIG. 10 exceptperformed with a different corn seed hybrid.

FIG. 12 shows a table with data from an experiment identical to theexperiment that generated the data for the table FIG. 10 exceptperformed with yet another corn seed hybrid.

FIG. 13 shows a table with data from an experiment conducted with barleyseed sonicated in argon saturated tap water.

FIG. 14 shows a table with data from a multiple trial experimentconducted on corn seed hybrid.

FIG. 15 shows a table with data from a multiple trial experimentconducted on corn seed hybrid.

FIG. 16 shows a table with data from a multiple trial experiment withthree different corn seed hybrids with various sonication times.

FIG. 17 shows a table and with data from an experiment identical to theexperiment that generated the data for the table of FIG. 16 performed at75% amplitude.

FIG. 18 shows the uptake of cresyl violet dye into corn seeds.

FIG. 19 shows a table with data from an experiment with toluidine bluestain in corn seeds.

FIG. 20 shows a barley seed.

FIG. 21 shows a table of data from an experiment conducted with cornseed hybrid sonicated in argon saturated tap water for various timeperiods.

FIG. 22 shows a table of data from several trial of an experimentconducted with corn seed hybrid saturated in argon saturated tap water.

FIG. 23 shows a table of data from an experiment identical to theexperiment that generated the data for the table of FIG. 22, exceptperformed for a different time period.

FIG. 24 shows a table of data from an experiment identical to theexperiment that generated the data for the table of FIG. 22, exceptperformed for a different time period.

FIG. 25 shows a table of data from an experiment identical to theexperiment that generated the data for the table of FIG. 22, exceptperformed for a different time period.

FIG. 26 shows a table of data from an experiment identical to theexperiment that generated the data for the table of FIG. 22, exceptperformed for a different time period.

FIG. 27 shows a table of data from an experiment identical to theexperiment that generated the data for the table of FIG. 22, exceptperformed for a different time period.

FIG. 28 shows an alternative apparatus for practicing the presentinvention.

DETAILED DESCRIPTION OF THE INVENTION

The invention describes a novel imbibition acceleration process 1) forthe uptake of a substance into a seed, particularly useful for enhancinga growth characteristic of the seed with that characteristictransferring to an advantage for the resultant plant, and 2) for theuptake of water for corn processing purposes. The growth characteristiccould be a resistance to a certain type of yield reducing pest, or aparticular growth advantage based on the introduction of a developmentalnutrient. In particular, pests can take the form of insects, weeds,mold, mildew and fungi. The substance responsible for imparting theenhanced growth characteristic could comprise an insecticide for givingthe seed a resistance to a particular variety of insects, a herbicidefor giving the seed a resistance to a particular variety of weed orplant, or a fungicide giving the seed a resistance to a particularvariety of mold, mildew or fungi.

In the way of further illustrative examples of applications of thepresent invention, it is anticipated that the present method isapplicable to corn wet milling used to produce corn starch, cornsweetener, corn oil, ethanol and animal feed by-products. Wet millingconsist, generally, of five main steps: 1) steeping; 2) germ separation;3) fiber washing and drying; 4) starch gluten separation; and 5) starchwashing. The steeping step involves soaking corn kernels in a solutionof sulfurous acid and water for a period of up to 48 hours. The steepingprocess cleans the corn kernels and softens the kernels to better allowthe cracking the kernels to remove the germ, which contains the oil. Asdemonstrated herein, the imbibition process of the present inventioncould dramatically reduce the time required for steeping by acceleratingthe uptake of a water solution into the corn kernel.

Moreover, imbibition plays a critical role in barley seed germination ofparticular interest in the field of factory malting. FIG. 20 shows abarley seed embryo 100, which includes a cotyledon 102 or the seed leaf,an epicotyl 104 which becomes the shoot, and a radicle 106 which becomesthe root. Additionally, FIG. 20 shows a seed coat 112, an endosperm 110,and the aleurone layer 108.

The malting process, used for the production of certain alcoholicbeverages, involves three basic steps: 1) steeping; 2) germination; and3) kilning. In barley steeping, the amount and the uniformity of wateruptake proves important. The time under water, the water temperature,the barley variety, and barley maturity comprise essential factors increating the correct cast moisture. Steeping takes place in steep tankswhere the barley seed 100 is mixed with a solution of water. The tanksare periodically roused with compressed air, to better ensure evenuptake of the steeping solution. A uniform moisture content is veryimportant to the quality and uniformity of the end product of themalting process.

The barley seeds 100 then begin to germinate. Barley's main role in themalting process is in the contribution of a rich source of sugar. Barleyseeds 100, however, in their dry state contain very little sugar, buthold a large reserve of starch in the endosperm 110. Starch is a polymerof sugar, and through an interaction during germination the starch isconverted to sugar. The biochemistry of this process begins with theimbibition of water through the seed coat 112 and into the interior ofthe barley seed. The water reacts with the cell embryo in a manner thatreleases a chemical known as gibberellic acid (GA), a plant hormone. TheGA is transported throughout the barley seed 100 until it arrives at thealeurone layer 108 that surrounds the endosperm 110. In the aleuronelayer 108, the GA acts to turn on certain genes in the nuclear DNA. Thegenes are transcribed resulting in the creation of messenger RNA, whichinteracts with a ribosome to begin the process of protein synthesis, ortranslation. The result is the creation of a protein called amylase. Theamylase is transported out from the aleurone cells 108 and into theendosperm 110. The amylase is an enzyme that acts as a catalyst for thehydrolysis of starch into sugar.

The process of converting the starch to sugar in barley seeds is dosedependent. In other words, the amount of GA present effects the rate anduniformity of the germination and conversion process. Consequently,imbibition of a solution of water and GA according to the methods of thepresent invention will significantly reduce the amount of time in themalting process, and will increase the rate and uniformity of thegermination and conversion of starch to sugar. Additionally, those ofordinary skill in the art will realize that the methods of the presentinvention apply equally to other growth hormones.

The imbibition process of the present invention is directed inparticular to such important agricultural seeds as corn, barley, andsoybeans by the sonication of such seeds in a liquid medium, preferablywater. Again, those of ordinary skill in the art will appreciate theapplicability of the present invention to other seeds types, withoutdeparting from the intended scope. The sonication is by the applicationof sound waves at ultrasonic frequencies from between about 15 kHz and100 kHz and preferably between about 20 kHz and 30 kHz, with an optimumnear 20 kHz.

Ultrasonic energy is applied to the liquid and seed mixture by a soundtransducer immersed in the liquid medium. While not wishing to be boundby any particular theory as to the mechanism of the subject of theinvention, it is currently believed that the acoustic energy is carriedthrough the liquid by oscillations of the liquid molecules in thedirection of propagation.

This produces alternating adiabatic compressions and decompressionstogether with corresponding increases and decreases in density andtemperature. If the periodic decreases of pressure in the liquid aresufficiently high during the negative pressure phase, the cohesiveforces of the liquid may be exceeded, at which point small cavities areformed by the process of cavitation. These small cavities then rapidlycollapse, producing a very large amplitude shock wave with localtemperatures up to a few hundred degrees centigrade or more. Thecollapse of the cavities are also known to create electrical dischargesupon their collapse, giving rise to the effect known assonoluminescence.

The effects of cavitation are greatly enhanced through the introductionof a variety of gases into the liquid. In the early 1930s, Frenzel andSchultes observed that photographic plates become exposed or fogged whensubmerged in water exposed to high frequency sound. This observation wasthe first recorded for the emission of light by acoustic waves orsonoluminescence. The physics of the phenomenon are not well understood.

With regard to the present invention, degassed distilled water requiresan energy density level of approximately 1 to 10 watts/cm² beforecavitation occurs. By saturating the water with a noble gas, such as oneor more of the inert gases helium, neon, argon, krypton, xenon, orradon, cavitation effects are seen at much lower energy density levelsand the effects at energy density levels on the order of 1 to 10watts/cm² are greatly enhanced. This effect is believed to be due to thecreation of microbubbles which more easily form the small cavities uponthe application of sonic energy. Additionally, the cavities in thepresence of the saturated gas are believed to generate shock waves oflarger amplitude upon collapse of the cavities than are achieved withdegassed water. In particular, it is believed that when tap water wassaturated with argon gas, helium gas, or argon and helium gasses,generally more dramatic uptake will be observed and such effects werereproducible from experiment to experiment. Other experiments in whichthe saturating gas was nitrogen also exhibited enhanced effects, but notnearly as pronounced as with argon. However, some experiments conductedwith tap water and with boiled double distilled water also producedsatisfactory results.

Since cavitation results in mechanical stress, sonication may create orenlarge fissures in the seed coat pericarp similar to scarification, awell-known process by which certain seeds, especially seeds with thickseed coats, are able to germinate. Scarification is believed toaccelerate imbibition of water through the pericarp. Simplescarification is unlikely to explain the novel effect disclosed herein,since scanning electron micrographs suggest no increase in the number offissures in treated seed, but do indicate a change in pericarp texture.It has been found that the sonication process accelerates the imbibitionof water. Cavitation may also result in physiological or biochemicalchanges in the seed which prime the germination process so that uponexposure of the seed to planting conditions, less time is needed for theseed to initiate germination, measured by the time when the radiclepushes through the pericarp. One mechanism proposed for causingphysiological or biochemical changes is the production of free radicalsby cavitaition.

The present method is carried out using an ultrasonic frequencygenerator for driving a piezoceramic sonicator, the horn of which isimmersed in the liquid surrounding the seeds. After sonication, theseeds are dried, and then placed on a water-saturated filter pad, or insome cases, in wet soil, to induce germination. The temperature duringgermination has been varied to analyze the effect of the treatment ongermination at various temperatures. Measurements which have beenmonitored in different experiments have included the time of emergenceof the primary root, the time of emergence of secondary roots, the timefor emergence of coleoptile, the root length and weight, the root area,the estimated volume of the root, the coleoptile length and weight, andthe uptake of water. The seeds tested were first generation (F₁) hybridseed corn.

Apparatus

The apparatus used in the treatment of seeds according to the presentinvention is illustrated diagrammatically in FIG. 1, generally at 10.Seeds 12 are placed in a container 14 and covered with a liquid medium16. A sound transducer 18 is suspended with the horn 20 of thetransducer immersed in the liquid medium 16. The transducer is connectedto an ultrasonic frequency generator 22. In the preferred embodiment,the sound transducer 18 is a piezoceramic transducer, Model VCX600obtained commercially from Sonics and Materials, Inc. Alternativetransducers may be used. Magnetostrictive transducers are capable ofdelivering higher levels of sound energy to the liquid media and may bepreferably used if higher sound densities are desired, for example iflarge quantities of seed are to be sonicated. The frequency generator 22is a Model 33120 Q obtained commercially from Hewlett Packard and ismatched to the transducer 18. It has a frequency range of between 15 kHzand 30 kHz and can supply between zero and 500 watts to the soundtransducer 18. In the experiments described herein, the power densitieswere between 30 watts per cm² and 80 watts per cm², although given therated efficiency of the sound transducer 18, higher power densities canbe achieved in the container 14.

FIG. 28 shows an alternative embodiment of an apparatus 100 of thepresent invention.

This differs from the apparatus 10 in the configuration of the cup horn130. The apparatus 100 includes an ultrasound frequency generator 122,and an acoustic actuator 118 (or sound transducer). These components aregenerally the same as the ultrasonic frequency generator 22, and thesound transducer 18 of apparatus 10. The cup horn 130 replaces the horn20 and container 14 of apparatus 10. In the apparatus 100 the cup horn130 comprises a single piece member, that includes a horn surrounded bya glass container. The horn of the cup horn 130 is generally longer andflatter than the horn 20 of apparatus 100 The cup horn 130 mountsupward, relative to its counterpart in apparatus 10. The sample restswithin the cup horn 130, otherwise, sonication takes place in a similarfashion regardless of the apparatus 10, 100 used. Water may becirculated through the wall of the cup horn 130 to maintain a constanttemperature.

Experiments

A series of experiments were performed to demonstrate the effectivenessof the methods of the present invention. FIG. 2 shows the results of anexperiment conducted with a Pioneer® #3394 corn seed hybrid. Theexperiment involved twenty trials with one seed per trial sonicated at20 kHz with a 3 mm probe, at an amplitude of 39%, for a period of 10minutes, in helium-saturated tap water. The seeds were placedindividually in a 14 ml test tube packed in ice. By comparison, 20Pioneer® #3394 corn seed hybrid seeds were soaked in helium-saturatedtap water for a period of 10 minutes. The weight in mg. of each of theseeds was measured prior to sonication and soaking, and measured againafter sonication and soaking. FIG. 2 shows the relative sonicated andsoaked weight difference in absolute amount, and in relative terms. Therelative percent water uptake reflects the weight gain as a percentageof the seed weight prior to sonication and soaking. The mean andstandard deviation across the entire experiment is reflected in the lastlines of the table depicted in FIG. 2, and shows clearly the enhanceduptake of water into the seeds due to the sonication process.

FIG. 3 shows the results of an experiment conducted with a Pioneer®#3939 corn seed hybrid. The experiment involved twenty trials with oneseed per trial sonicated at 20 kHz with a 3 mm probe, at an amplitude of39%, for a period of 10 minutes, in argon saturated tap water. The seedswere placed individually in a 14 ml test tube packed in ice. Bycomparison, 20 Pioneer® #3939 corn seed hybrid seeds were soaked inargon saturated tap water for a period of 10 minutes. The weights in mg.of each of the seeds was measured prior to sonication and soaking, andmeasured again after sonication and soaking. FIG. 3 shows the relativesonicated and soaked weight difference in absolute amount, and inrelative terms. The relative percent water uptake reflects the weightgain as a percentage of the seed weight prior to sonication and soaking.The mean and standard deviation across the entire experiment isreflected in the last lines of the table depicted in FIG. 3, and againshows clearly the enhanced uptake of water into the seeds due to thesonication process.

FIG. 4 repeats the experiment described above and represented by thedate shown in FIG. 3, except with Pioneer® #3963 corn seed hybrid seeds.Similarly FIG. 4 shows that superior water uptake result from thesonication process.

FIG. 5 repeats the experiment described above and represented by thedate shown in FIG. 2, except that argon and helium saturated tap waterwas used for both the sonicated and soaked groups, again with similarresults.

FIG. 6 repeats the experiment described above and represented by thedata shown in FIG. 3, except with Pioneer® #5005 sweet corn hybridseeds.

FIG. 7 repeats the experiment described above and represented by thedata shown in FIG. 2, except that argon saturated tap water was used forboth the sonicated and the soaked groups.

FIG. 8 repeats the experiment described above and represented by thedata shown in FIG. 2, except that tap water was used for both thesonicated and the soaked groups.

FIG. 9 repeats the experiment described above and represented by thedata shown in FIG. 2, except that boiled double distilled water was usedfor both the sonicated and the soaked groups.

FIG. 10 shows the results of an experiment conducted with a Pioneer®#3394 corn seed hybrid. The experiment involved a trial with 20 seedsper trials sonicated at 20 kHz with a 45 mm probe, at an amplitude of30%, for a period of 10 minutes, in argon saturated tap water. Twelvegroups of 20 seeds each were placed in a 2″ diameter aluminum cup packedin ice. By comparison, twelve groups of 20 Pioneer® #3394 corn seedhybrid seeds were soaked in argon saturated tap water for a period of 10minutes. The weight of 20 seeds in mg. was measured prior to sonicationand soaked and the average weight of seed determined. They were measuredagain after sonication and soaking. FIG. 10 shows the average weight perseed for each of the 12 groups of 20 seeds. FIG. 10 shows the relativesonicated and soaked weight difference in absolute amount, and inrelative terms. The relative percent water uptake reflects the weightgain as a percentage of the seed weight prior to sonication and soaking.The mean and standard deviation across the entire experiment isreflected in the last lines of the table depicted in FIG. 10, and againshows clearly the enhanced uptake of water into the seeds due to thesonication process.

FIG. 11 repeats the experiment described above and represented by thedata shown in FIG. 10, except that Pioneer® #3820 corn seed hybrid wasused.

FIG. 12 repeats the experiment described above and represented by thedata shown in FIG. 10, except that Pioneer® #3963 corn seed hybrid wasused.

FIG. 13 shows the results of an experiment conducted with a No. 3-141barley seed, provided by Briess Malting Company of Chilton, Wis. Theexperiment involved twenty trials with one seed per trial sonicated at20 kHz with a 3 mm probe, at an amplitude of 39%, for a period of 8minutes, in argon saturated tap water. By comparison, twenty No. 3-141barley seeds were soaked in argon saturated tap water for a period of 8minutes. The weights in mg. of each of the seeds was measured prior tosonication and soaking, and measured again after sonication and soaking.FIG. 13 shows the average weight per seed for each of the twenty seeds.

FIG. 13 shows the relative sonicated and soaked weight difference inabsolute amount, and in relative terms. The relative percent wateruptake reflects the weight gain as a percentage of the seed weight priorto sonication and soaking. The mean and standard deviation across theentire experiment is reflected in the last lines of the table depictedin FIG. 11, and the results show that the barley seeds react to thesonication process in a manner similar to the corn hybrid seeds.

FIG. 14 shows the results of a series of experiments conducted with aPioneer® #9281 soybean seed hybrid. The experiment involved three groupseach comprised of five trials, each trial in turn comprising twentyseeds sonicated at 20 kHz with a 45 mm probe, at an amplitude of 30%, inargon saturated tap water. Group 1 was sonicated for 2 minutes, group 2was sonicated for 4 minutes, and group 3 was sonicated for 6 minutes.Each groups of twenty seeds were placed in a 2″ diameter aluminum cuppacked in ice. By comparison, the soaking portion of the experimentswere performed on three groups each comprising five trials, each trialcomprising twenty Pioneer® #9281 soybean hybrid seeds soaked in argonsaturated tap water. Group 1 was soaked for 2 minutes, groups 2 wassoaked for 4 minutes and group 3 was soaked for 6 minutes. The weight inmg. of each of the seeds was measured prior to sonication and soaking,and measured again after sonication and soaking. FIG. 14 shows theweight for the entire seed groups of the five trials of twenty seeds foreach of the three groups. FIG. 14 shows the relative sonicated andsoaked weight difference in absolute amount, and in relative terms. Therelative percent water uptake reflects the weight gain as a percentageof the seed weight prior to sonication and soaking. The mean andstandard deviation across the entire experiment is reflected in the lastlines of each groups as shown in FIG. 14, and again shows clearly theenhanced uptake of water into the seeds due to the sonication process.

FIG. 15 shows the results of a series of experiments conducted with thePioneer® #3394 corn seed hybrid. The experiments involved several trialsof twenty seeds per trial sonicated at 20 kHz with a 45 mm probe, at anamplitude of 39%, in argon saturated tap water. The sonication timevaried from 2 minutes to 12 minutes in 2 minute increments. Each groupof twenty seeds were placed in a 2″ diameter aluminum cup packed in ice.By comparison, the soaking portion of the experiments was performed ongroups of twenty Pioneer® #3394 corn hybrid seeds soaked in argonsaturated tap water. The soaking time varied from 2 minutes to 12minutes in 2 minute increments. FIG. 15 shows the total weight for thetwenty seed groups for each of the varying sonication and soaking timegroups. FIG. 15 shows the relative sonicated and soaked weightdifference in absolute amount, and in relative terms for the entire seedgroups.

The relative percent water uptake reflects the weight gain as apercentage of the seed weight prior sonication and soaking. The resultsallow comparison of the relative amounts of water uptake for sonicationand for soaking, and show that over all time period involved sonicationproduces superior results.

FIG. 16 shows the results of a series of experiments conducted withthree different hybrid corn seeds, Pioneer® #3394, Pioneer® #3573, andPioneer® #65672 Honey & Pearl respectively. The experiment involvedseveral trials of twenty seeds per trial sonicated at 20 kHz with a 13mm probe, at an amplitude of 100%, in argon saturated tap water. Thesonication time varied from 2 minutes to 10 minutes in 2 minuteincrements. Each groups of twenty seeds were placed in a 125 ml glassbeaker packed in ice. By comparison, the soaking portion of theexperiments was performed on one group of twenty seeds for each of theabove-identified hybrids. The soaking time was fixed at 10 minutes foreach of the hybrids. FIG. 16 shows the total weight for the twenty seedgroups for each of the varying hybrids, sonication time groups, andsoaking groups. FIG. 16 shows the relative sonicated and soaked weightdifference in absolute amount, and in relative terms for the entire seedgroups. The relative percent water uptake reflects the weight gain as apercentage of the seed weight prior to sonication and soaking. Theresults allow for comparison of the relative amounts of water uptake forsonication over a variety of time periods for three different hybrids.In all cases, the uptake percentages for the sonicated seeds exceededthe uptake from soaking for the same hybrids.

FIG. 17 repeats the experiment described above and represented by thedata shown in FIG. 16, except that an amplitude of 75% was used.

FIG. 18 shows that sonication promotes the uptake of a cresyl violet dyeinto corn seeds. Sonication facilitates the uptake of the dye into thecorn seeds, and into the embryo of the seed. By contrast the controlseeds show that soaking accomplishes only minimal uptake, which does notpenetrate beyond the periphery of the seed. The results depicted in FIG.18 demonstrate the effectiveness of the sonication method in introducingsubstances of differing molecular weights into seeds.

In a similar manner, FIG. 19 shows that the sonication process is alsoeffective at introducing a toluidine blue stain into corn seeds. In thisset of experiments sonication took place in two groups, with five trialsper group, for a total time of 15 minutes per trial. The sonicationsolution contained 15 ml of water and 750 mg. of toluidine blue stain.After each trial the amount of stain absorbed was determined. After thefive trials in each of the groups the total stain absorbed wascalculated, and a percentage of stain uptake was calculated based on theamount of stain absorbed over the five trials as a percentage of thetotal amount of stain originally present. The procedure was repeated fortwo soaked groups each comprising five trials. FIG. 18 shows that thesonication process results in dramatically higher absolute and relativeamounts of uptake of the toluidine blue stain.

The experiments described below were conducted with the apparatus 100,rather than the apparatus 10 used above. FIG. 21 shows the results of anexperiment conducted with Pioneer® #3394 corn seed hybrid. Theexperiment involved six groups of one seed each soncicated for 1, 2, 4,6, 8, and 10 minutes in tap water saturated with 5 ml. of argon, at 30%amplitude in a polypropylene tube suspended in a cup horn 130 with coldwater circulation. Each of the six groups represents the results oftwenty individual trials. In comparison, the same experiment wasperformed for the same time groups, twenty trials each, except that theseeds were soaked only. The weights in mg. were measured prior tosonication and soaking, and again afterwards. FIG. 21 shows the totalweights for each of the different groups of twenty seeds, and therelative and absolute weight differences for the sonicated and soakedgroups. The results of this experiment show that the apparatus 10, 100produce similar results, and that the sonicated seeds exhibit superioruptake of the solution when compared to the soaked seeds.

FIG. 22 shows the results of an experiment conducted with Pioneer® #3394corn seed hybrid. The experiment involved twenty trials with one seedper trial sonicated at 20 kHz, at an amplitude of 39%, for a period of 1minute, in argon saturated tap water. The seeds were placed individuallyin a polypropylene tube suspended in a large cup horn 130 with coldwater circulation. By comparison, twenty Pioneer® #3394 corn hybridseeds were soaked in argon-saturated tap water for a period of 1 minute.The weight in mg. of each of the seeds was measured prior to sonicationand soaking, and measured again after sonication and soaking. FIG. 22shows the relative sonicated and soaked weight difference in absoluteamount, and in relative terms. The relative percent water uptakereflects the weight gain as a percentage of the seed weight prior tosonication and soaking. The mean and standard deviation across theentire experiment is reflected in the last lines of the table depictedin FIG. 22, and shows clearly the enhanced uptake of water into theseeds due to the sonication process.

FIG. 23 shows the results of an experiment identical to the experimentdescribed above and represented by the data shown in FIG. 22, exceptthat the time was extended from 1 minute to 2 minutes.

FIG. 24 shows the results of an experiment identical to the experimentdescribed above and represented by the data shown in FIG. 22, exceptthat the time was extended from 1 minute to 4 minutes.

FIG. 25 shows the results of an experiment identical to the experimentdescribed above and represented by the data shown in FIG. 22, exceptthat the time was extended from 1 minute to 6 minutes.

FIG. 26 shows the results of an experiment identical to the experimentdescribed above and represented by the data shown in FIG. 22, exceptthat the time was extended from 1 minute to 8 minutes.

FIG. 27 shows the results of an experiment identical to the experimentdescribed above and represented by the data shown in FIG. 22, exceptthat the time was extended from 1 minute to 10 minutes.

The series of experiments depicted in FIGS. 22-27 demonstrates theresponse to uptake over time for sonicated and soaked groups of seed,and again demonstrates the superiority of the sonication process. Inprinciple, the overall impact of the above experiments demonstrates theability of the imbibition process to uptake a variety of substances intoseeds of a variety of different plants. As a result, introduction of anynumber of substances capable of imparting growth enhancingcharacteristics to a plant can be affected through the imbibitionprocess of the present invention.

As is demonstrated in more detail in the commonly owned co-pending U.S.patent application Ser No. 08/886,901 filed Jul. 2, 1997 entitled METHODFOR ENHANCING GERMINATION, incorporated herein by reference, theenhanced growth characteristic should continue to effect the seed andresultant plant for extended periods of time. Enhanced germinationeffects continue to manifest in sonicated seeds after periods ofextended drying down and rehydration.

Although the invention has been described with respect to a preferredembodiment thereof, it is to be also understood that it is not to besole limited since changes and modifications can be made therein whichare within the full intended scope of this invention as defined by theappended claims.

We claim:
 1. An imbibition process for the uptake of a substance into aseed, comprising the steps of: a) immersing said seed in a liquidsolution that includes a dissolved gas and a pesticide capable ofenhancing a growth characteristic of said seed; b) introducing into saidliquid sound energy at a frequency and energy density sufficient tocreate cavitation in said liquid; and c) sonicating said seed for aperiod of time sufficient to result in an improved rate of uptake ofsaid pesticide into said seed.
 2. The invention in accordance with claim1 wherein said pesticide comprises an insecticide.
 3. The invention inaccordance with claim 1 wherein said pesticide comprises a herbicide. 4.The invention in accordance with claim 1 wherein said pesticidecomprises a fungicide.